Analysis of samples and determination of the concentration of components contained therein is a common and important process in chemistry and biology. Of particular importance is the analysis of biological fluids such as blood, urine or saliva to determine the concentrations of various components. Also of great importance is the measurement of the concentration of various chemical species embedded within a biological material such as tissue.
The chemical analysis of blood, urine, and other biological fluids is crucial to the diagnosis, management, treatment and cure of a wide variety of diseases and medical conditions including diabetes, kidney disease and heart disease. In the case of diabetes, monitoring of blood glucose levels several times per day is a necessary feature of management of the disease for many patients. In the case of people with diseases of the circulatory system, the analysis of various blood components is of importance both in diagnosis and in treatment. For example, the level of cholesterol compounds of various types in the blood of man has a strong correlation with the probability of onset of atherosclerosis. In patients with renal insufficiencies, urine analysis provides valuable information relating to kidney function. In a related application, the concentration of alcohol in blood is known to be correlated to an individual's physical response times and can provide information relating to, for example, the individuals fitness to drive a motorized vehicle.
At present, analysis of biological fluid for these and other applications is commonly invasively performed, that is by removing a sample of fluid, and subjecting it to one or more chemical tests. Typically, separate tests are required for each analyte to be measured. These tests require the use of consumable supplies and reagents and are moderately expensive. Often skilled technicians are needed to remove the fluid, and to perform the chemical tests. Frequently the tests are made in centralized clinical laboratories with resulting complexity of sample tracking, and quality control. In such circumstances there are additional problems relating to the potential change in the chemical composition of the fluid between its extraction and its analysis. Furthermore the turnaround time for such measurements can be undesirably long.
For many applications it would be desirable to be able to make real-time measurements of analytes in biological fluids or of the level of various clinically important chemicals in tissue. Ideally these measurements would be made non-invasively.
In addition to the above applications, there are a number of instances in which it is desirable to measure the local concentration of chemical species in tissue either in-vivo or in-vitro. In the case of victims of stroke or of head trauma it is desirable to be able to monitor the degree of brain edema as well as the concentration of various metabolic chemical species in the brain that act as indicators of brain function. These include various fatty acid compounds, water, blood, lactates, and various proteins and lipids. Other specific examples include the monitoring of metabolic function by measurement of tissue oxygenation, or the measurement of localized changes in tissue blood perfusion such as may be indicative of hyperplastic or neoplastic tissue.
Furthermore, it is widely believed that the ability to monitor certain changes in tissue chemical composition may lead to predictive tests for various types of cancer. Examples of such changes are the development of microcalcifications, specific changes in tissue chromophore types and concentrations, and specific variation in tissue hormone levels. Consequently non-invasive methods and apparatus which enabled measurements to be made of the chemical composition of tissue samples in-vivo would also be a very important development.
At present, when it is desired to monitor chemical levels in tissue such as in the brain, existing techniques require the use of Magnetic Resonance Imaging, CAT scan imaging or PET scan imaging. All of these techniques require the use of expensive equipment and do not allow bedside or continuous monitoring. Furthermore, for many conditions even these expensive and complex analytical techniques do not supply adequate information about the concentration of specific chemical species in the tissue.
An as yet unrealized goal for in-vivo monitoring of biological fluids and of the chemical composition of tissue would therefore be the development of methods and apparatus for the non-invasive, real-time measurement of analytes in a cost effective manner. For in-vitro fluid analysis the ability to make rapid measurements of single or multiple analytes could decrease analysis times, thus boosting the throughput of the clinical laboratories and reducing the cost of the analyses.
One approach to non-invasively determining the composition of tissue or of a biological fluid makes use of the interaction of electromagnetic radiation with the matter under examination. It is known that electromagnetic radiation having appropriate characteristics may interact with matter in two primary ways. As it passes through the material the radiation will be scattered and a portion of it will be absorbed. Different chemical species scatter and absorb to different degrees at different wavelengths. The physical composition of the medium will also effect its interaction with the radiation. A number of methods have been proposed that use optical radiation to probe tissue or fluid samples with the goal of determining the concentration of a component of the material by making use of known characteristics of the relationship between optical absorption of the medium and wavelength.
These prior methods generally share a number of common elements. A source of optical radiation emits light which enters the medium of interest and interacts with the medium, with the result that some radiation is absorbed by the medium. The incident light is chosen so that it contains wavelengths that are partially or wholly absorbed by the species for which the concentration is to be measured. Subsequently, the transmitted, reflected or scattered light is detected, and its intensity as a function of wavelength is measured. The measured intensity spectrum is then analyzed in order to yield information relating to the concentration of chemical species of interest within the medium.
Such prior art is represented, for example, by the development of the pulsed oximeter such as is described in U.S. Pat. No. 4,621,643 (New, Jr. et al., 1986). Such a device allows the determination of the percentage of oxygen saturation of the blood (i.e. the relative saturation).
In a series of patents, typified by U.S. Pat. No. 4,223,680 (Jobsis, 1980) and U.S. Pat. No. 4,281,645 (Jobsis, 1981), Jobsis has described the use of similar optical intensity measurements, to measure relative tissue oxygenation and metabolism by making use of the characteristic optical absorption spectra of both haemoglobin and of the cellular enzyme cytochrome (a,a.sub.3). As with the work of New, the invention of Jobsis utilizes a small number of discrete wavelengths of light.
In extensions of this work, various techniques have been proposed in which many wavelengths of optical radiation would be used to measure the concentration of multiple analytes within a biological fluid or within biological tissue. After propagating into the medium and interacting with the component of interest the reflected or transmitted optical radiation is detected and the concentration of the component of interest is determined by comparison of the wavelength dependent intensity spectrum of the detected light with the known absorption spectra of a family of target components.
Such prior art is represented by U.S. Pat No. 4,975,581 (Robinson et al., 1990). An important aspect of such art is the use of sophisticated statistical techniques for the analysis of the measured optical absorption spectra. The study of these techniques is the science of chemometrics and the details of chemometric analysis are widely described in the scientific literature.
It is known to those skilled in the art of chemometrics that the number of independent chemical species that can be analyzed in a medium by the optical absorption spectroscopy techniques described above can not exceed the number of independent optical wavelengths used in the analysis. This is a fundamental limitation of the early inventions of New and of Jobsis. Since they rely on the use of a limited small number of discrete wavelengths, they can only be used in the case that optical absorption is primarily caused by a small number of chemical species.
This is the case for the measurement of oxygen in blood. However for most other chemical species of interest in biological media, this is not the case since there are numerous chemical species with similar and overlapping absorption spectra. In such instances the use of many wavelengths and multivariate analysis, such as is discussed in the work of Robinson et al, in U.S. Pat. No. 4,975,581, is essential.
All of the above prior art shares a fundamental flaw which makes its use for the analysis of chemical species in biological media suboptimal. The parameter measured by these techniques is the attenuation of light as it passes through a medium such as tissue or blood. From this attenuation is deduced the optical absorption of the medium. The optical absorption at a given wavelength is proportional to the absorption per molecule of chemical species times the concentration of the chemical times the distance travelled by the light in the medium. If we know the absorption properties of the chemical and we know the distance travelled by the light in the tissue, the individual chemical concentration can be deduced.
In biological media, and in fact in any scattering material, light is scattered multiple times as it traverses the medium. As a result, the path length travelled by the light is considerably longer than the direct geometric distance from light source to light detector. In fact, in typical tissue such as brain tissue, the light may travel 4 to 6 times that distance. Furthermore the distance travelled by the light will depend on the wavelength of the light and on the scattering and absorption properties of the medium at that wavelength. Consequently, in many cases optical spectroscopic techniques are able to provide only information about the optical absorption of a material and hence about the chemical concentration of a particular chemical species integrated over the path length travelled by the light in the medium. The reduction of that information to an absolute measurement of chemical concentration requires knowledge of the path length travelled in the medium.
This knowledge is not provided by any of the prior art techniques. Instead, they rely on the use of approximate estimates of path length, or measure parameters such as ratios of concentrations in which path length cancels. As a result, there are many important clinical situations in which it is desirable to obtain absolute concentrations of chemical species for which the prior art techniques are inadequate or insufficiently accurate. In this invention is described a method and apparatus for determining absolute chemical concentrations in highly scattering media by combining optical spectroscopic analysis with simultaneous path length analysis of the measured light.